Investigators have linked the cases to burgers eaten at the SSE Hydro, a major entertainment venue which opened in Glasgow in September last year.
They identified three days between Friday January 17 and Sunday January 19 as the likely infection period.
During this time, the venue hosted five performances of the popular motoring show, Top Gear Live.
The £125 million Hydro has a capacity of 12,000, meaning up to 36,000 people could have attended its events during that weekend.
Previously the days up until January 25 - which included a performance by Del Amitri at Celtic Connections - had been investigated, but NHS Greater Glasgow and Clyde (NHSGGC) confirmed the dates of the infection period had been narrowed to the events of the weekend.
Seven cases of the bug were first reported at the end of January, with the number steadily rising since.
All but one of those affected by E.coli live in Scotland, with one living in Cumbria, the health board.
It added that all 21 people affected were either fully recovered or were recovering at home.
Three people from the Greater Glasgow area contracted the infection, three from Lanarkshire, and five from the Lothian area.
Two cases were people from the Highlands, four from the Forth Valley, two from the Grampian region, and one from Fife.
Dr Eleanor Anderson, consultant in public health medicine with NHSGGC said: "We are pleased that all the cases are recovering at home.
"While there is no conclusive evidence, our investigations have indicated that the most likely source is a link to the consumption of burgers at the SSE Hydro.
"Glasgow City Council environmental health officers have worked closely with the vendors and will continue to liaise with them to ensure all appropriate food hygiene standards are being met."
The bug, specifically E.coli O157, is a harmful strain of the bacteria which can cause serious illness.
It is spread through eating or drinking contaminated food or drink, or contact with infected animals or people. Symptoms include stomach cramps, diarrhoea, nausea and fever.
The health board said that, as the incubation period for anyone who attended the Hydro between January 17 and January 19 has now passed, it was unlikely that any more people would develop symptoms.
However, they said that they may still continue to identify people who have already been unwell.
They also urged anyone who was at the Hydro during that specific weekend and experienced any of the symptoms to contact their GP so all cases could be recorded.
A spokeswoman for the Hydro said they have been contacted by environmental health officials and were "awaiting further details to establish the exact cause of this isolated incident".
She added: "The initial review of our caterers' standard procedures have been found satisfactory and Environmental Health have seen nothing in their processes which concerns them."
The next big concert at the entertainment venue is singer Beyonce, who is set to perform two shows on February 20 and February 21.
It is also set to host gymnastics and netball events in the Commonwealth Games in July.